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Effects of Sex and Autism on Oxytocin Receptors in the Substantia Nigra of the Human Brain

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Effects of Sex and Autism on Oxytocin Receptors in the Substantia Nigra of the Human Brain Utah State University DigitalCommons@USU Undergraduate Honors Capstone Projects Honors Program 5-2021 Effects of Sex and Autism on Oxytocin Receptors in the Substantia Nigra of the Human Brain Kip Dooley Utah State University Follow this and additional works at: https://digitalcommons.usu.edu/honors Part of the Biology Commons Recommended Citation Dooley, Kip, "Effects of Sex and Autism on Oxytocin Receptors in the Substantia Nigra of the Human Brain" (2021). Undergraduate Honors Capstone Projects. 692. https://digitalcommons.usu.edu/honors/692 This Thesis is brought to you for free and open access by the Honors Program at DigitalCommons@USU. It has been accepted for inclusion in Undergraduate Honors Capstone Projects by an authorized administrator of DigitalCommons@USU. For more information, please contact [email protected]. EFFECTS OF SEX AND AUTISM ON OXYTOCIN RECEPTORS IN THE SUBSTANTIA NIGRA OF THE HUMAN BRAIN By Kip Dooley Capstone submitted in partial fulfillment of the requirements for graduation with UNIVERSITY HONORS With a major in Human Biology in the department of Biology Approved: ______________________________ _______________________________ Capstone Mentor Departmental Honors Advisor Dr. Sara Freeman Dr. Brett Adams ___________________________________ Director of University Honors Program Dr. Kristine Miller UTAH STATE UNIVERSITY Logan, UT Spring 2021 © 2021 Kip Dooley All Rights Reserved i Abstract Oxytocin, a hormone present in the mammalian brain, has been shown to be a vital component of social function in animals and may have a role in the social deficits associated with Autism Spectrum Disorder in humans. Based on previous studies from our lab, there are oxytocin receptors in the human substantia nigra, a basal ganglia structure in the midbrain that is important in both movement and reward pathways. The substantia nigra contains two subsections that are defined by the neurotransmitters they contain: the pars compacta, which is dopaminergic, and the pars reticulata, which is GABAergic. By localizing oxytocin receptors in either the pars compacta or pars reticulata, we can infer the role of that region as it relates to social function. We previously attempted to identify the pars compacta using immunohistochemistry for tyrosine hydroxylase, but the background signal was too high to reliably be used to delineate the boundaries, so we are trying a new approach. We used Nissl staining, which has been shown to reveal dopaminergic neurons in the substantia nigra and has been used to distinguish the pars compacta from the pars reticulata. Once identified, we used the borders of the pars compacta to quantify oxytocin receptors within the substantia nigra in a neuroanatomically informed way by overlaying microscope images of tissue (with the pars compacta outlined) with the receptor autoradiographs, which visualize oxytocin receptors in the substantia nigra. The tissue was acquired from four distinct groups: eight typically developing (TD) males, seven TD females, eight males with Autism Spectrum Disorder, and seven females with Autism Spectrum Disorder. We analyzed the oxytocin receptor binding to determine the effect of sex and autism on oxytocin receptor density in the pars compacta. Females with ASD exhibited significantly reduced OXTR density when compared to both males with ASD and TD females, which may be related to differences in expression of symptoms between males with ASD and females with ASD. Future ii directions of this research are aimed at defining the role of the oxytocin system in individuals with Autism Spectrum Disorder and how it relates to the social deficits present in those individuals. iii Acknowledgements First, I would like to thank my research and capstone mentor, Dr. Sara Freeman for the support and guidance she has offered throughout both the capstone process and my time in her lab. I am very grateful to have had the opportunity to be a part of her lab and contribute to the amazing work she does. Her abundant knowledge and willingness to guide me through the writing process has been invaluable to my capstone project as well as my academic writing abilities, and for that I can not thank her enough. I would also like to acknowledge Michelle Palumbo, who sliced and mounted the brain tissue used in this project. Lastly, I want to thank my incredible support system in my friends and especially my family. My family, particularly my mom, has been incredibly supportive throughout this process and was very willing to listen and offer support in conversations about my capstone project. Table of Contents Introduction ................................................................................................................................... 1 Materials and Methods ................................................................................................................. 3 Results ............................................................................................................................................ 7 Discussion..................................................................................................................................... 10 Reflective Writing ....................................................................................................................... 14 References .................................................................................................................................... 17 Professional Author Bio ............................................................................................................. 19 List of Figures Figure 1: Total, nonspecific, and specific OXTR autoradiograms ....................................... 6 Figure 2: Nissl-stained section and specific binding autoradiogram .................................... 6 Figure 3: Effect of sex and ASD on OXTR density in the pars compacta of the SN .......... 8 Figure 4: Association between age and OXTR density in the SN ......................................... 8 Figure 5: Association between postmortem interval and OXTR density in the SN ............ 9 Figure 6: Association between ASD symptom severity and OXTR density ...................... 10 1 Introduction Oxytocin (OT) has been shown to play an important role in social function in animals and is known to modulate social behavior (Carter et al., 2020). Because of its relationship with social functions, OT is implicated in the biology of Autism Spectrum Disorder (ASD), which is a developmental neuropsychiatric condition that is characterized in part by social deficits. OT is a neurotransmitter that acts in the brain by binding to the OT receptor (OXTR). Examining the site of action of OT can provide insight into its function as it relates to ASD in both males and females. Analyses of OXTR density differences between TD and ASD specimens may reveal some of the mechanisms of ASD and its associated social deficits. ASD is a complicated condition that is a major area of ongoing research and is characterized by three classes of symptoms. These include difficulty interacting socially, deficits in both verbal and non-verbal communication, and repetitive or harmful behavior patterns (Beuker et al., 2012). ASD predominantly affects males, who are four times as likely to be diagnosed with ASD than females, which may be due to neuroanatomical differences or underdiagnosis in females (Fombonne, 2009). In order to gain a better understanding of ASD, many studies, including this one, focus on differences in brain chemistry and anatomy between typically developing brain tissue and brain tissue from individuals with ASD. The substantia nigra (SN) is a small structure in the midbrain that contains two anatomically interwoven yet functionally distinct subunits, the dopaminergic pars compacta and the GABAergic pars reticulata. The pars compacta is involved in reward pathways and fine motor control while the pars reticulata has an inhibitory effect on downstream neural targets when stimulated (Caputi et al., 2013; see also Hodge & Butcher, 1980). Previously, Paval (2017) proposed that autistic behavior is partially a result of abnormalities in the dopaminergic system 2 housed in the midbrain. Previous work from our lab has identified dense OXTR binding in the SN of the human brain but has not been able to identify which of these two subunits the signal resides in. Knowing the specific location of the OXTR within the SN and whether they are present in one or both subdivisions influences how we interpret the function of OT in that area. Other studies, such as Jacob et al. (2007), provide support for a connection between the OXTR gene and ASD in certain populations. Ribeiro et al. (2018) identified two polymorphisms at the OXTR gene that may play a role in the diagnosis of ASD and its associated behaviors. Over a decade of clinical research has provided evidence for symptom improvement in ASD after treatment with intranasal OT, although some studies have reported no changes (Guastella et al., 2015 see also; Anagnostou et al., 2012). However, for an OT treatment to be effective, the OXTR system must be working properly. Freeman et al. (2018) found evidence for dysregulated OXTR in the ASD brain. However, like the other studies mentioned previously, this study did not evaluate sex differences or study the SN. While these studies have suggested that dysfunctions and differences in both dopamine and OT circuits are potential contributing factors to ASD, neuroanatomical support for these hypotheses is lacking. Our
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